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. 2015 Nov 10;10(11):e0141231. doi: 10.1371/journal.pone.0141231

Fig 3. Clodronate liposome mediated macrophage depletion in Col4a3KO mice.

Fig 3

(A) KO male mice were dosed intraperitonally with CL (KO+CL) or PBSL (KO+PBSL) as control. Animals were entered in the study at the age of 4 weeks and were continually dosed until the age of 8 weeks. Injections were repeated every second day, except for the first two doses, which were injected on consecutive days. (B-D) CL significantly reduced F4/80 positive macrophage infiltrates in KO+CL kidney as compared to KO+PBS or KO mice at the age of 8 weeks. (B, C) Representative images and quantitative assessment of F4/80 stained macrophages in kidney sections from KO+CL and KO+PBSL reveal marked reduction in macrophage infiltrates following CL dosing. Scale bar: 100 μm. (D) Real-time PCR analysis of F4/80 mRNA expression in KO+CL, KO+PBSL, KO, and WT littermates at the age of 8 weeks. Significant reduction in F4/80 mRNA expression following CL dosing was observed. PBSL did not affect endogenous F4/80 mRNA expression as shown by similar F4/80 mRNA expression in KO and KO+PBSL mice. (E) Real-time PCR analysis of CD204 and CD206 mRNA showed significant reduction in the expression of both genes in KO kidney upon CL treatment. (F) Representative images and quantitative assessment of F4/80 and CD206 stained kidney sections from KO+CL and KO+PBSL revealed marked reduction in M1 (F4/80+CD206-) and M2 (F4/80+CD206+) macrophages. Asterisk: F4/80+CD206- M1 macrophages, arrow: F4/80+CD206+ M2 macrophages. Scale bar: 20 μm. Welch corrected ANOVA followed by posthoc pairwise t-test with a Bonferroni correction (C,D,E) and unpaired t-test (F) was used to evaluate differences between groups. (C,D,E,F) n = 5 mice per group. CL: clodronate liposomes; PBSL: PBS liposomes.